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A novel persulfide detection method reveals protein persulfide- and polysulfide-reducing functions of thioredoxin and glutathione systems.
Sci Adv. 2016 Jan; 2(1):e1500968.SA

Abstract

Hydrogen sulfide signaling involves persulfide formation at specific protein Cys residues. However, overcoming current methodological challenges in persulfide detection and elucidation of Cys regeneration mechanisms from persulfides are prerequisites for constructing a bona fide signaling model. We here establish a novel, highly specific protein persulfide detection protocol, ProPerDP, with which we quantify 1.52 ± 0.6 and 11.6 ± 6.9 μg/mg protein steady-state protein persulfide concentrations in human embryonic kidney 293 (HEK293) cells and mouse liver, respectively. Upon treatment with polysulfides, HEK293 and A549 cells exhibited increased protein persulfidation. Deletion of the sulfide-producing cystathionine-γ-lyase or cystathionine-β-synthase enzymes in yeast diminished protein persulfide levels, thereby corroborating their involvement in protein persulfidation processes. We here establish that thioredoxin (Trx) and glutathione (GSH) systems can independently catalyze reductions of inorganic polysulfides and protein persulfides. Increased endogenous persulfide levels and protein persulfidation following polysulfide treatment in thioredoxin reductase-1 (TrxR1) or thioredoxin-related protein of 14 kDa (TRP14) knockdown HEK293 cells indicated that these enzymes constitute a potent regeneration system of Cys residues from persulfides in a cellular context. Furthermore, TrxR1-deficient cells were less viable upon treatment with toxic amounts of polysulfides compared to control cells. Emphasizing the dominant role of cytosolic disulfide reduction systems in maintaining sulfane sulfur homeostasis in vivo, protein persulfide levels were markedly elevated in mouse livers where hepatocytes lack both TrxR1 and glutathione reductase (TR/GR-null). The different persulfide patterns observed in wild-type, GR-null, and TR/GR-null livers suggest distinct roles for the Trx and GSH systems in regulating subsets of protein persulfides and thereby fine-tuning sulfide signaling pathways.

Authors+Show Affiliations

Department of Molecular Immunology and Toxicology, National Institute of Oncology, Ráth György utca 7-9, Budapest 1122, Hungary.Division of Biochemistry, Department of Medical Biochemistry and Biophysics, Karolinska Institutet, SE-171 77 Stockholm, Sweden.Department of Molecular Immunology and Toxicology, National Institute of Oncology, Ráth György utca 7-9, Budapest 1122, Hungary.Division of Biochemistry, Department of Medical Biochemistry and Biophysics, Karolinska Institutet, SE-171 77 Stockholm, Sweden.Division of Biochemistry, Department of Medical Biochemistry and Biophysics, Karolinska Institutet, SE-171 77 Stockholm, Sweden.Department of Molecular Immunology and Toxicology, National Institute of Oncology, Ráth György utca 7-9, Budapest 1122, Hungary.Department of Microbiology and Immunology, Montana State University, Cooley Hall, PO Box 173520, Bozeman, MT 59717, USA.Division of Redox Regulation, German Cancer Research Center (DKFZ), DKFZ-ZMBH Alliance, Im Neuenheimer Feld 280, 69120 Heidelberg, Germany.Division of Redox Regulation, German Cancer Research Center (DKFZ), DKFZ-ZMBH Alliance, Im Neuenheimer Feld 280, 69120 Heidelberg, Germany.Department of Microbiology and Immunology, Montana State University, Cooley Hall, PO Box 173520, Bozeman, MT 59717, USA.Division of Biochemistry, Department of Medical Biochemistry and Biophysics, Karolinska Institutet, SE-171 77 Stockholm, Sweden.Department of Molecular Immunology and Toxicology, National Institute of Oncology, Ráth György utca 7-9, Budapest 1122, Hungary.

Pub Type(s)

Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't

Language

eng

PubMed ID

26844296

Citation

Dóka, Éva, et al. "A Novel Persulfide Detection Method Reveals Protein Persulfide- and Polysulfide-reducing Functions of Thioredoxin and Glutathione Systems." Science Advances, vol. 2, no. 1, 2016, pp. e1500968.
Dóka É, Pader I, Bíró A, et al. A novel persulfide detection method reveals protein persulfide- and polysulfide-reducing functions of thioredoxin and glutathione systems. Sci Adv. 2016;2(1):e1500968.
Dóka, É., Pader, I., Bíró, A., Johansson, K., Cheng, Q., Ballagó, K., Prigge, J. R., Pastor-Flores, D., Dick, T. P., Schmidt, E. E., Arnér, E. S., & Nagy, P. (2016). A novel persulfide detection method reveals protein persulfide- and polysulfide-reducing functions of thioredoxin and glutathione systems. Science Advances, 2(1), e1500968. https://doi.org/10.1126/sciadv.1500968
Dóka É, et al. A Novel Persulfide Detection Method Reveals Protein Persulfide- and Polysulfide-reducing Functions of Thioredoxin and Glutathione Systems. Sci Adv. 2016;2(1):e1500968. PubMed PMID: 26844296.
* Article titles in AMA citation format should be in sentence-case
TY - JOUR T1 - A novel persulfide detection method reveals protein persulfide- and polysulfide-reducing functions of thioredoxin and glutathione systems. AU - Dóka,Éva, AU - Pader,Irina, AU - Bíró,Adrienn, AU - Johansson,Katarina, AU - Cheng,Qing, AU - Ballagó,Krisztina, AU - Prigge,Justin R, AU - Pastor-Flores,Daniel, AU - Dick,Tobias P, AU - Schmidt,Edward E, AU - Arnér,Elias S J, AU - Nagy,Péter, Y1 - 2016/01/22/ PY - 2015/07/20/received PY - 2015/11/20/accepted PY - 2016/2/5/entrez PY - 2016/2/5/pubmed PY - 2016/2/5/medline KW - Glutathione system KW - Life sciences KW - ProPerDP method KW - Proteins KW - Thioredoxins KW - biochemistry KW - cell signaling KW - hydrogen sulfide KW - persulfide SP - e1500968 EP - e1500968 JF - Science advances JO - Sci Adv VL - 2 IS - 1 N2 - Hydrogen sulfide signaling involves persulfide formation at specific protein Cys residues. However, overcoming current methodological challenges in persulfide detection and elucidation of Cys regeneration mechanisms from persulfides are prerequisites for constructing a bona fide signaling model. We here establish a novel, highly specific protein persulfide detection protocol, ProPerDP, with which we quantify 1.52 ± 0.6 and 11.6 ± 6.9 μg/mg protein steady-state protein persulfide concentrations in human embryonic kidney 293 (HEK293) cells and mouse liver, respectively. Upon treatment with polysulfides, HEK293 and A549 cells exhibited increased protein persulfidation. Deletion of the sulfide-producing cystathionine-γ-lyase or cystathionine-β-synthase enzymes in yeast diminished protein persulfide levels, thereby corroborating their involvement in protein persulfidation processes. We here establish that thioredoxin (Trx) and glutathione (GSH) systems can independently catalyze reductions of inorganic polysulfides and protein persulfides. Increased endogenous persulfide levels and protein persulfidation following polysulfide treatment in thioredoxin reductase-1 (TrxR1) or thioredoxin-related protein of 14 kDa (TRP14) knockdown HEK293 cells indicated that these enzymes constitute a potent regeneration system of Cys residues from persulfides in a cellular context. Furthermore, TrxR1-deficient cells were less viable upon treatment with toxic amounts of polysulfides compared to control cells. Emphasizing the dominant role of cytosolic disulfide reduction systems in maintaining sulfane sulfur homeostasis in vivo, protein persulfide levels were markedly elevated in mouse livers where hepatocytes lack both TrxR1 and glutathione reductase (TR/GR-null). The different persulfide patterns observed in wild-type, GR-null, and TR/GR-null livers suggest distinct roles for the Trx and GSH systems in regulating subsets of protein persulfides and thereby fine-tuning sulfide signaling pathways. SN - 2375-2548 UR - https://www.unboundmedicine.com/medline/citation/26844296/A_novel_persulfide_detection_method_reveals_protein_persulfide__and_polysulfide_reducing_functions_of_thioredoxin_and_glutathione_systems_ L2 - https://doi.org/10.1126/sciadv.1500968 DB - PRIME DP - Unbound Medicine ER -